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A new diagnostic test for COVID-19 based on CRISPR technology can be run on nasal swabs.

CRISPR has come to COVID-19.

The US Food and Drug Administration has granted Sherlock Biosciences an emergency use authorization (EUA) for its COVID-19 diagnostic assay, beating out other companies and academic groups trying to use the powerful gene-editing technology to figure out who is infected with the novel coronavirus.

The EUA, which makes Sherlock’s test the first FDA-authorized use of CRISPR technology for anything, allows the company to scale up production of its assay for use by laboratories that do complex diagnostics.

CEO Rahul Dhanda says the test will be inexpensive and can be done in about an hour. He says the EUA represents the maturing of a technology that holds an incredible amount of potential in understanding disease.

Sherlock’s test is a molecular diagnostic, intended to identify people who have acute SARS-CoV-2 infection. It capitalizes on a CRISPR-based technology developed in the lab of Feng Zhang, a scientist at Broad Institute of MIT and Harvard and a cofounder of Sherlock.

The test enters a crowded field of SARS-CoV-2 diagnostics. Although Dhanda says Sherlock’s test can be run on basic machines that many hospitals likely have, it remains to be seen who decides to vet this new technology against standard tests.

Like those standard tests, Sherlock’s assay detects the presence of the viral RNA. It starts with a respiratory specimen from, for example, the mouth, nose, or lungs. To make the viral genome easier to identify, scientists convert it into DNA, which can be copied over and over. The method they use—isothermal amplification—is done at a constant temperature, unlike the method used by most conventional diagnostics, polymerase chain reaction.

Then, the sample goes through Sherlock’s CRISPR gauntlet. CRISPR-Cas is a bacterial defense system that chops up invasive viral RNA; scientists have turned it into a technique that makes precise cuts in genetic code through various Cas enzymes. Sherlock’s system uses Cas13, which William Blake, the firm’s chief technology officer, says is a little more flexible in what genetic regions it can target than other Cas enzymes.

The CRISPR part of the assay involves converting the amplified DNA back into RNA, which is the type of genetic information the Cas13 enzyme recognizes. The enzyme is led to any viral RNA in the sample based on “guides,” short bits of RNA scientists add to the reaction that match the actual code of the virus. Once it’s there, the enzyme cuts the viral RNA.

Blake says the assay targets two distinct parts of the SARS-CoV-2 genome: the recipe for the nucleocapsid, which helps the virus assemble itself, and ORF 1ab, a stretch of the genome that leads to the precursor of an enzyme that helps the virus copy itself.

Blake says those targets were chosen over better known ones like the SARS-CoV-2 spike protein and the protease because viruses are a bit sloppy when they copy their genetic information. Sometimes they make mistakes, and while those mistakes may not affect the virus’ ability to copy itself and infect, it might affect the precision of a diagnostic based on CRISPR.

“We wanted to ensure that our tests enabled detection of all the sequences that are out there for SARS-CoV-2,” he says.

Once it’s activated, the Cas13 enzyme cuts other nucleic acids as well as the viral RNA, Blake says. And that nonspecific cutting is how Sherlock knows the reaction has worked. Within the assay are strands of genetic material that have a fluorescent molecule at one end, and a molecule that quenches, or blocks, the fluorescence, on the other. As activated Cas13 chomps its way around the genetic material in the sample, it cuts those strands, freeing the fluorescents bits from the quenching bits. Blake says that most fluorescent plate readers can read the test.

This is different from how the technology is being used as a gene-editing device, Blake says, where it has to be very specific and have no extraneous cutting.

Like other SARS-CoV-2 diagnostics to receive EUAs, Sherlock’s test was validated through experiments to determine analytical sensitivity, its ability to work on clinical samples, and tests to determine cross reactivity, Dhanda says. That validation was conducted using samples from people who had tested negative for SARS-CoV-2, or who had a non-COVID-19 respiratory disease. The diagnostic has not been tested in a hospital setting yet.

Dhanda says the company is working with an experienced manufacturer and will be able to produce tens of thousands of tests per week. Its main customers for now, he says, will be hospitals, a setting that should provide real-world data about the fidelity of the first CRISPR-based diagnostic.

“When it is authorized, and launched, we’ll have real-world scenarios to really compare it, but today, we have very high confidence in that performance being more than necessary to diagnose COVID-19,” Dhanda says.

Mammoth Biosciences, a CRISPR diagnostic company cofounded by Jennifer Doudna of the University of California, Berkeley, is trying to push its own COVID-19 test to market. Chief Technology Officer Janice Chen tells C&EN that Mammoth has filed its EUA application with the FDA for its test.

Earlier this year, the Mammoth team published a peer-reviewed study of the system and the effort to validate it, as some molecular diagnostic tests came under fire for their poor reliability (Nat. Biotechnol. 2020 DOI: 10.1038/s41587-020-0513-4).

Like Sherlock’s assay, Mammoth’s test starts with a constant-temperature amplification step, but its CRISPR-Cas system looks for different viral targets, including the envelope protein of SARS-CoV-2. Their readout can be done two ways: with fluorescence, similarly to the Sherlock product, or via lateral flow—adding the final processed sample to a small cassette and looking for a color change signal that is similar to a pregnancy test. Sherlock is also working on a lateral-flow system.

Caspr Biotech is also working on a CRISPR COVID-19 diagnostic. In early March, the firm published proof-of-principle details of its assay to a preprint server—where it has yet to undergo peer review—also analyzing the use of lateral flow as a readout (bioRxiv 2020, DOI: 10.1101/2020.02.29.971127).

In all three cases, the companies were working on CRISPR-based diagnostics for other diseases, and as information about SARS-CoV-2 grew, they decided to shift large portions of their efforts to the growing pandemic. While these tests may not end up widely used, COVID-19 has presented an opportunity to get CRISPR technology, and the platforms upon which diagnostics can be built, into hospitals much faster than under normal circumstances.

In March, as Mammoth was beginning to navigate San Francisco’s shelter-in-place order and validating its test using patient samples from UCSF, Chen described to C&EN why the pivot to SARS-CoV-2 was important.

“Based on everything we are living through today, having this tool available would be hugely beneficial to public health efforts,” she said. “What we’ve learned with this public health crisis is that there’s actually also a large need for getting point-of-care patient testing available. In some ways, this is kind of an important milestone.”